Each year in the United States there are close to 30,000 hospitalizations in specialized burn centers. Many of these patients benefit from human skin allografts as temporary wound closure to promote rapid healing. Skin allografts, being highly immunogenic, begin to lose viability shortly after transplantation and are completely rejected by cytotoxic T cells within two weeks. Extending skin allograft survival is an unmet medical need because it will prolong the duration in which burn victims are protected from dehydration and infection. Regulatory T cells (Treg) have the potential to prolong the viability of skin allografts, but adoptive cell therapy of Treg is not feasible in urgent care settings. Systemic infusion of the principal Treg inhibitory molecules, namely TGF-?, IL-10, and CTLA-4 (TIC), is complicated by off-target toxicities and the different pharmacokinetics of the agents. To address this need, we seek to develop a versatile platform on which TIC are optimized spatiotemporally for prolonging the survival of skin allografts. In this project we propose to deliver TIC simultaneously and locally into skin allografts established on full-MHC mismatched mice. The Treg mimicking strategy entails formulating TIC into an injectable by intermixing with novel bioaffinity amphiphilic peptides, generating a gel in which Fc-fusion proteins can be loaded. The multivalent, multifunctional (?multiplexing?) system will enforce a powerful multi-pronged immunosuppression at the transplant site by cross-linking TGF-? and IL-10 receptors, as well as B7 molecules on target leukocytes. Three specific aims will be carried out to delineate the cellular and molecular mechanisms of the strategy. The first task is to optimize the dose combination of TIC with respect to raising Treg to Th1 ratio in an allogeneic skin explant T cell co-culture system. In the second task, we will validate the biocompatibility of the materials system. In the third task, we will evaluate the capacity of muxTIC to steer T cells toward suppressive phenotypes in vivo. The public health impact centers on leveraging an enabling technology by which burn care can be improved. Given the in vivo transplant model at hand and the biomaterials tools we have developed, our team is uniquely poised to advance the novel strategy. The materials platform is versatile; Fc-fusion proteins or antibodies targeting other pathways can be displayed, thereby increasing the scope of the modulation. The broader impact is that the data generated will set the stage for testing multiplexed Treg factors in vascularized composite allografts (VCA) transplantation in which the skin is a primary driver of immune rejection.